In the United States and throughout the world, cancer incidence and mortality has increased dramatically in both developed and developing nations. Cancer causes ~13% of human deaths with 7.6 million people dying from cancer in 2007. More people in the US die of lung cancer than breast, colon, kidney, and prostate cancers combined. Recent studies show that veterans are 25 to 75 percent more likely to develop lung cancer than people who did not serve in the military; yet therapies for lung cancer and other solid tumors are still limited. Recent successes in T cell cancer immunotherapy point to a potential breakthrough in treatment. T cells expressing chimeric antigen receptors or tumor-reactive ?? TCRs have cured patients with advanced metastatic disease. Intrinsic T cell immunity against tumors can be released using mAbs to remove inhibition by checkpoint CTLA-4 and PD-1 receptors which has resulted in a number of cures in melanoma and lung cancer. Yet, significant limitations exist for these therapies. Therapy is limited to certain cancers, not all patients respond to therapy, and there is significant toxicity. Although showing great promise, additional approaches to cancer immunotherapy are needed. Treatment with ?? T cells expressing V?2V?2 TCRs is one such therapy that shows promise. In contrast to ?? T cells, the antigen responses of ?? T cells expressing V?2V?2 TCRs are not MHC restricted. The major subset of human ?? T cells use their V?2V?2 T cell receptors to recognize the foreign-microbial isoprenoid metabolite, HMBPP, and the self-metabolite, IPP. Normal cells and tumor cells from a wide variety of tissues can stimulate V?2V?2 cells. V?2V?2 T cells expand to very high numbers during many infections (up to 1 in 2 circulating T cells) and can kill tumor cells and infected cells as well as secrete inflammatory cytokines, chemokines, and growth factors. Two approaches are being used to treat cancer with V?2V?2 T cells. The first is to immunize with stimulators such as the bromohydrin analog of HMBPP or the aminobisphosphonate, zoledronate, with low-dose IL-2. The second is to adoptively transfer V?2V?2 T cells grown ex vivo. This approach has cured a patient with metastatic kidney cancer, induced remission in another with breast cancer, and induced partial remissions or stable disease in other patients but needs to be made more effective. Metabolic engineering of bacteria is a new field of study that has focused on altering bacteria for drug or chemical synthesis. Changes in bacterial metabolism are made by modifying biochemical pathways or by introducing new ones. We have now provided proof-of-principle for this approach by metabolic engineering Salmonella to overproduce HMBPP and demonstrating responses in monkeys. We now propose to improve our Salmonella vaccine and to test a new Listeria vaccine. Both species have been used for cancer vaccines but differ significantly because Salmonella is given orally whereas Listeria is given intravenously. We will use the bacterial vaccines to target and activate adoptively transferred V?2V? 2 T cell in tumors because they preferentially localize to tumor cells. To accomplish our goals, we will: metabolically engineer bacteria to overproduce HMBPP, test engineered bacteria in vitro and in vivo in monkeys, and assess the ability of metabolically engineered bacteria to target and activate adoptively transferred V?2V?2 T cells to control tumors. We have an outstanding team with an excellent track record and have extensive experience working with ?? T cells and isoprenoid metabolism. We have established in vivo models and techniques. A proof-of-principle Salmonella vaccine has been derived and the results recently published. The molecular methods to create more vaccines are well developed. In conclusion, immunotherapy using metabolically engineered bacterial vaccines with V?2V?2 T cells has the potential to be broadly applicable for the treatment of many different tumors both by direct activation and through adoptive transfer.